It has been suggested to use O.sup.6 -alkyl guanine derivatives possessing O.sup.6 -alkylguanine-DNA alkyltransferase depleting activity in order to enhance the effectiveness of chemotherapeutic alkylating agents used for killing tumour cells. There is increasing evidence that in mammalian cells the toxic and mutagenic effects of alkylating agents are to a large extent a consequence of alkylation at the O.sup.6 -position of guanine in DNA. The repair of O.sup.6 -alkylguanine is mediated by ATase, a repair protein that acts on the O.sup.6 -alkylated guanine residues by stoichiometric transfer of the alkyl group to a cysteine residue at the active site of the repair protein in an autoinactivating process. The importance of ATase in protecting cells against the biological effects of alkylating agents has been most clearly demonstrated by the transfer and expression of cloned ATase genes or cDNAs into ATase deficient cells; this confers resistance to a variety of agents, principally those that methylate or chloroethylate DNA. Whilst the mechanism of cell killing by O.sup.6 -methylguanine in ATase deficient cells is not yet clear, killing by O.sup.6 -chloroethylguanine occurs through DNA interstrand crosslink formation to a cytosine residue on the opposite strand via a cyclic ethanoguanine intermediate, a process that is prevented by ATase-mediated chloroethyl group removal or complex formation.
The use of O.sup.6 -methylguanine and O.sup.6 -n-butylguanine for depleting ATase activity has been investigated (Dolan et al., Cancer Res., (1986) 46, pp. 4500; Dolan et al., Cancer Chemother. Pharmacol., (1989) 25. pp 103. O.sup.6 -benzylguanine derivatives have been proposed for depleting ATase activity in order to render ATase expressing cells more susceptible to the cytotoxic effects of chloroethylating agents (Moschel et al., J. Med.Chem., 1992, 35, 4486). U.S. Pat. No. 5,091,430 and International Patent Application No. WO 91/13898 Moschel et al. disclose a method for depleting levels of O.sup.6 -alkylguanine-DNA alkyl-transferase in tumour cells in a host which comprises administering to the host an effective amount of a composition containing O.sup.6 -benzylated guanine derivatives of the following formula: ##STR2## wherein Z is hydrogen, or ##STR3## and R.sup.a is a benzyl group or a substituted benzyl group. A benzyl group may be substituted at the ortho, meta or para position with a substituent group such as halogen, nitro, aryl such as phenyl or substituted phenyl, alkyl of 1-4 carbon atoms, alkoxy of 1-4 carbon atoms, alkenyl of up to 4 carbon atoms, alkynyl of up to 4 carbon atoms, amino, monoalkylamino, dialkylamino, trifluoromethyl, hydroxy, hydroxymethyl, and SO.sub.n R.sup.b wherein n is 0, 1, 2 or 3 and R.sup.b is hydrogen, alkyl of 1-4 carbon atoms or aryl. Mi-Young Chae et al, J.Med.Chem., 1994, 37, 342-347 describes tests on O.sup.6 -benzylguanine analogs bearing increasingly bulky substituent groups on the benzene ring or at position 9. Mi-Young Chae et. al., J. Med. Chem. 1995, 38, 359-365 published on 20 Jan. 1995 describe several 8-substituted O.sup.6 -benzylguanines, 2- and/or 8-substituted 6-(benzyloxy)purines, substituted 6(4)-(benzyloxy)pyrimidines, and a 6-(benzyloxy)-s-triazine which were tested for their ability to inactivate ATase . Two types of compounds were identified as being significantly more effective than O.sup.6 -benzylguanine at activating ATase in human HT29 colon tumour cell extracts. These were 8-substituted O.sup.6 -benzylguanines bearing electron-withdrawing groups at the 8-position (e.g. 8-aza-O.sup.6 -benzylguanine and O.sup.6 -benzyl-8-bromoguanine) and 5-substituted 2,4-diamino-6-(benzyloxy)pyrimidines bearing electron withdrawing groups at the 5-position (e.g. 2,4-diamino 6-(benzyloxy)-5-nitroso- and 2,4-diamino-6-(benzyloxy)-5-nitropyrimidine). The latter derivatives were also more effective than O.sup.6 -benzylguanine at inactivating ATase in intact HT29 colon tumour cells.
The present inventors are also inventors in U.S. patent application Ser. No. 08/568,576, entitled "O.sup.6 -substituted guanine derivatives, a process for their preparation and use thereof in treating tumour cells" filed 7 Dec. 1995 as a Continuation-In-Part of International Application PCT/IE94/00031 which has an international filing date of 8 Jun. 1994 and which was published under No. WO 94/29312 on 22 Dec. 1994. The U.S. Application and WO 94/29312 (the contents of which are incorporated herein by reference in their entirety) describe O.sup.6 -substituted guanine derivatives of formula I: ##STR4## wherein
Y is H, ribosyl, deoxyribosyl, or R"XCHR'", wherein X is O or S, R" and R'" are alkyl, or substituted derivatives thereof;
R' is H, alkyl or hydroxyalkyl;
R is (i) a cyclic group having at least one 5- or 6-membered heterocyclic ring, optionally with a carbocyclic or heterocyclic ring fused thereto, the or each heterocyclic ring having at least one hetero atom chosen from O, N, or S, or a substituted derivative thereof; or
(ii) naphthyl or a substituted derivative thereof; PA1 a) if R.sup.2 and R.sup.5 are NH.sub.2 and R.sup.4 is NO or NO.sub.2 PA1 b) if R.sup.2 is NH.sub.2 and R.sup.4 and R.sup.5 form a ring structure X ##STR7## c) if R.sup.2 is NH.sub.2 and R.sup.4 and R.sup.5 for a ring structure XI ##STR8## d) if R.sup.2 is NH.sub.2, R.sup.4 is NO.sub.2 and R.sup.5 is H or CH.sub.3 PA1 e) if R.sup.2, R.sup.4 and R.sup.5 are NH.sub.2. PA1 f) if R.sup.2 and R.sup.5 are NH.sub.2 and R.sup.4 is H PA1 g) if R.sup.2 is H, R.sup.4 is NO.sub.2 and R.sup.5 is NH.sub.2 PA1 h) if R.sup.2 is F or OH, and R.sup.4 and R.sup.5 form a ring structure XII ##STR9##
and pharmaceutically acceptable salts thereof.
It is an object of the present invention to provide novel compounds useful for depleting ATase activity in order to enhance the effects of chemotherapeutic agents such as chloroethylating or methylating anti-tumour agents. It is a further object to provide compounds having better ATase inactivating characteristics than O.sup.6 -benzylguanine and having different solubility patterns.
Another object of the invention is to provide pharmaceutical compositions containing compounds which are useful for depleting ATase activity. A further object of the present invention is to provide a method for depleting ATase activity in tumour cells. A still further object of the invention is to provide a method for treating tumour cells in a host.
The present invention provides 6-hetarylalkyloxy pyrimidine derivatives of formula II ##STR5## wherein
R is as defined in (i) above, or R is phenyl or a substituted derivative thereof,
R.sup.2 is selected from H, C.sub.1 -C.sub.5 alkyl, halogen or NH.sub.2, R.sup.4 and R.sup.5 which are the same or different are selected from H, NH.sub.2 or NO.sub.n where n 1 or 2, or R.sup.4 and R.sup.5 together with the pyrimidine ring form a 5- or 6-membered ring structure containing one or more hetero atoms, and pharmaceutically acceptable salts thereof,
provided that R.sup.2 is not NH.sub.2 if R.sup.4 and R.sup.5 form a ring structure IX ##STR6##
wherein Y is H, ribosyl, deoxyribosyl, arabinosyl, or R"XCHR'" wherein X is O or S, R"" and R'" are alkyl, or subsituted derivatives thereof, and provided that R is not phenyl;
R may suitably be a 5- or 6-membered heterocyclic ring or a benzo derivative thereof, in which latter case the 6-alkyloxypyrimidine moiety may be attached to R at either the heterocyclic or the benzene ring.
In preferred embodiments, R is a 5-membered ring containing S or O, with or without a second ring fused thereto.
Preferably, R is a heterocyclic ring having at least one S atom; more preferably, R is a 5-membered heterocyclic ring having at least one S atom; and most preferably, R is a thiophene ring or a substituted derivative thereof.
Alternatively, R may be a heterocyclic ring having at least one O atom, particularly, a 5-membered heterocyclic ring having at least one O atom and more particularly R may be a furan ring or a substituted derivative thereof.
As another alternative, R may be a heterocyclic ring having at least one N atom, particularly R may be a 6-membered heterocyclic ring having at least one N atom and in particular, R may be a pyridine ring.
In the definition of Y, the term "substituted derivative" includes substitution by one or more of the following groups: hydroxy, halo, alkoxy, amino, alkylamino, amido or ureido.
In the definition of R, the term "substituted derivative" includes substitution of the heterocyclic rings and/or carbocyclic ring(s) by one or more of the following groups: alkyl, alkenyl, alkynyl, halo, haloalkyl, nitro, cyano, hydroxyalkyl, SO.sub.n R"" where R"" is alkyl and n=0,1 or 2, or a carboxyl or ester group of the formula --COOR.sup.5 wherein R.sup.5 is H or alkyl. Halo, haloalkyl, cyano, alkylenedioxy SO.sub.n R"" (as defined above) and --COOR.sup.5 wherein R.sup.5 is alkyl are preferred substituents.
An alkyl, alkenyl, or alkynyl group preferably contains from 1 to 20, more preferably from 1 to 10 and most preferably from 1 to 5 carbon atoms. Halo includes iodo, bromo, chloro or fluoro.
One embodiment of the invention provides a pharmaceutical composition containing compounds of formula II, wherein Y, R, R.sup.2, R.sup.4 and R.sup.5 are as defined above, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient. Optionally the composition may also contain an alkylating agent such as a chloroethylating or methylating agent.
In a further embodiment, the present invention provides a method for depleting Abase activity in a host comprising administering to the host an effective amount of a composition containing a compound of formula II wherein Y, R, R.sup.2, R.sup.4 and R.sup.5 are as defined above, or a pharmaceutically acceptable salt thereof, more particularly a pharmaceutical composition as defined above. This method may alternatively be defined as a method of depleting ATase mediated DNA repair activity in a host.
The invention further provides a method for treating tumour cells in a host comprising administering to the host an effective amount of a composition containing a compound of formula II wherein Y, R, R.sup.2, R.sup.4 and R.sup.5 are as defined above or a pharmaceutically acceptable salt thereof, more particularly a pharmaceutical composition as defined above and administering to the host an effective amount of a composition containing an alkylating agent. The method may be used for treatment of neoplasms including those which are known to be sensitive to the action of alkylating agents e.g. melanoma and glioma and others whose resistance to treatment with alkylating agents alone may be overcome by the use of an inactivator according to the invention.
Subject to the provisos above the preferred compounds of the invention are those of: ##STR10## wherein:
R is as defined for formula II, particularly furyl or thienyl unsubstituted or substituted, preferahly with a halogen such as chlorine, bromine or fluorine,
Y is H or HOCH.sub.2 CH.sub.2 OCH.sub.2 -;
R.sup.2 is H, NH.sub.2, C.sub.1 -C.sub.5 alkyl, preferably methyl, or halogen, preferably fluorine;
R.sup.3 is H or OH; ##STR11## wherein:
R is as defined for formula II, particularly phenyl, thienyl or furyl unsubstituted or substituted preferably with a halogen such as chlorine, bromine or fluorine, or phenyl having a methylenedioxy ring structure fused thereto;
Y is as defined for formula II;
X is CH or N;
A is CH or N; and preferably when X=N, A=CH ##STR12## wherein:
R is as defined for formula II
X is CH or N
A is CH or N; ##STR13## wherein:
R is as defined for formula II, particularly phenyl, thienyl or furyl unsubstituted or substituted preferably with a halogen such as chlorine or bromine;
Z is O or S or CH.dbd.CH; ##STR14## wherein:
R is as defined for formula II;
U is CH or N;
V is CH or N;
W is CH or N; ##STR15## wherein:
R is as defined for formula II, particularly phenyl optionally substituted with halogen preferably one or more of chlorine, fluorine or bromine, or methylenedioxy; thenyl or furyl optionally substituted with halogen preferably one or more of chlorine, bromine or fluorine;
T is H, NH.sub.2 or NO.sub.n where n-=1 or 2;
Q is H, NH.sub.2 or NO.sub.n where n=1 or 2;